Limitations of the widely used GAM42a and BET42a probes targeting bacteria in the Gammaproteobacteria radiation

Yeates, C, Saunders, AM, Crocetti, GR and Blackall, LL (2003) Limitations of the widely used GAM42a and BET42a probes targeting bacteria in the Gammaproteobacteria radiation. Microbiology-sgm, 149 5: 1239-1247. doi:10.1099/mic.0.26112-0


Author Yeates, C
Saunders, AM
Crocetti, GR
Blackall, LL
Title Limitations of the widely used GAM42a and BET42a probes targeting bacteria in the Gammaproteobacteria radiation
Journal name Microbiology-sgm   Check publisher's open access policy
ISSN 1358-0872
Publication date 2003-01-01
Sub-type Article (original research)
DOI 10.1099/mic.0.26112-0
Volume 149
Issue 5
Start page 1239
End page 1247
Total pages 9
Place of publication Great Britain
Publisher Society for General Microbiology
Language eng
Subject C1
270307 Microbial Ecology
780105 Biological sciences
270301 Bacteriology
710702 Waste management and recycling
Abstract The 23S rRNA-targeted probes GAM42a and BET42a provided equivocal results with the uncultured gammaproteobacterium 'Candidatus Competibacter phosphatis' where some cells bound GAM42a and other cells bound BET42a in fluorescence in situ hybridization (FISH) experiments. Probes GAM42a and BET42a span positions 1027-1043 in the 23S rRNAand differ from each other by one nucleotide at position 1033. Clone libraries were prepared from PCR products spanning the 16S rRNA genes, intergenic spacer region and 23S rRNA genes from two mixed cultures enriched in 'Candidatus C. phosphatis'. With individual clone inserts, the 16S rDNA portion was used to confirm the source organism as 'Candidatus C. phosphatis' and the 23S rDNA portion was used to determine the sequence of the GAM42a/BET42a probe target region. Of the 19 clones sequenced, 8 had the GAM42a probe target (T at position 1033) and 11 had G at position 1033, the only mismatch with GAM42a. However, none of the clones had the BET42a probe target (A at 1033). Non-canonical base-pairing between the 23S rRNA of 'Candidatus C. phosphatis' with G at position 1033 and GAM42a (G-A) or BET42a (G-T) is likely to explain the probing anomalies. A probe (GAM42_C1033) was optimized for use in FISH, targeting cells with G at position 1033, and was found to highlight not only some 'Candidatus C. phosphatis' cells, but also other bacteria. This demonstrates that there are bacteria in addition to 'Candidatus C. phosphatis' with the GAM42_C1033 probe target and not the BET42a or GAM42a probe target.
Keyword Microbiology
Water Treatment Processes
In-situ Detection
Activated-sludge
Ribosomal-rna
Accumulating Organisms
Identification
Removal
Diversity
Proteobacteria
Cells
Q-Index Code C1

Document type: Journal Article
Sub-type: Article (original research)
Collections: 2004 Higher Education Research Data Collection
Advanced Water Management Centre Publications
 
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Created: Wed, 15 Aug 2007, 12:20:38 EST